Nα-2-(4-nitrophenylsulfonyl)ethoxycarbonyl-amino acids

ABSTRACT

Protected amino acid derivatives of general formula (I)                    
     and methods for the prepatation of the derivatives are provided.

This application is the national phase under 35 U.S.C. §371 of PCTInternational Application No. PCT/KR96/00179 which has an Internationalfiling date of Oct. 19, 1996 which designated the United States ofAmerica.

BACKGROUND OF THE INVENTION

The field of the invention concerns new protected amino acid derivativesuseful for peptide synthesis, namely,N_(α)-2-(4-nitrophenylsulfonyl)ethoxycarbonyl-amino acids having thegeneral formula I:

wherein R₁ represents a hydrogen atom, and R₂ may representhydroxymethyl, 1-hydroxyethyl, 4-hydroxybenzyl, imidazolyl-2-methyl,benzyloxymethyl, 1-benzyloxyethyl, 4-benzyloxybenzyl,benzyloxycarbonylmethyl, 2-(benzyloxycarbonyl)ethyl, S-benzylthiomethyl,S-(diphenylmethyl)thiomethyl, 4-(benzyloxycarbamido)butyl,3-guanidinopropyl, 3-N^(G)-toluenesulfonyl) guanidinopropyl,3-(N^(G)-nitro) guanidinopropyl,3-[N^(G)-(4-methoxy-2,3,6-trimethylbenzenesulfonyl)]-guanidinopropyl,N-(4,4′-dimethoxybenzhydryl)carboxamidomethyl,2-[N-(4,4′-dimethoxybenzhydryl) carboxamido]ethyl,S-tert-butyldithiomethyl,4-[2-(4-nitrophenylsulfonyl)ethoxycarbamicdo]-butyl, propyl, butyl,ethyl, 3-(benzyloxycarbamido)propyl, or 3-(tert-butoxycarbamido)propyl,and methods for the preparation thereof.

N₆₀ -2-(4-Nitrophenylsulfonyl) ethoxycarbonyl-amino acids (Nsc-aminoacids) I represent a class of protected amino acid derivatives which areused in the chemical synthesis of peptides. In these derivatives,N_(α)-2-(4-nitrophenylsulfonyl) -ethoxycarbonyl (Nsc) group serves as atemporary N_(α)-protection which can be selectively removed after eachstep of the peptide chain elongation. Nsc-Group is fairly resistant toacidic reagents and is cleaved according to the β-elimination mechanismby organic bases in aprotic solvents. Mild conditions of the cleavageallow for use of the temporary N_(α)-Nsc-protection in the peptidesynthesis together with the acid-sensitive side chain protection ofwidely used t-butyl or benzyl type, thus providing the so-called“orthogonality” of the synthetic strategy.

Recently N_(α)-Nsc-amino acids, methods for their preparation, and theiremployment for the solid phase peptids synthesis have been disclosed inInternational Publication No. WO96/25394. However, only a minimal set ofN_(α)-Nsc-derivatives of proteogenic amino acids has been described,which is intended for the solid phase peptide synthesis with fullside-chain protection of acid-labile tert-butyl or compatible type.

SUMMARY OF THE INVENTION

It is, therefore, desirable to develop new N_(α)-Nsc-amino acids toextend the list of derivatives available for solid phase synthesis. Onthe other hand, it is useful to provide N_(α)-Nsc-amino acid derivativeswith another type of side protection (e.g., benzyl of compatible) orwithout side protection which may give new opportunities for liquidphase peptide synthesis.

An object of the present invention is to provide new N_(α)-Nsc-aminoacids derivatives, more particularly,N_(α)-2-(nitrophenylsulfonyl)ethoxycarbonyl-amino acids having thegeneral formula I:

wherein R₁ represents hydrogen atom, and R₂ may represent ahydroxymethyl, 1-hydroxyethyl, 4-hydroxybenzyl, imidazolyl-2-methyl,benzyloxymethyl, 1-benzyloxyethyl, 4-benzyloxybenzyl,benzyloxycarbonylmethyl, 2- (benzyloxycarbonyl) ethyl,S-benzylthiomethyl, S-(diphenylmethyl)thiomethyl,4-(benzyloxycarbamido)butyl, 3-guanidinopropyl, 3-(N^(G)-toluenesulfonyl) guanidinopropyl, 3-(N^(G)-nitro)guanidinopropyl,3-[N^(G)-(4-methoxy-2,3,6-trimethylbenzenesulfonyl)]-guanidinopropyl,N-(4,4′dimethoxybenzhydryl) carboxamidomethyl,2-[N-(4,4′-dimethoxybenzhydryl)carboxamido]ethyl,S-tert-butyldithiomethyl,4-[(2-(4-nitrophenylsulfonyl)ethoxycarbamido]-butyl, propyl, butyl,ethyl, 3-(benzyloxycarbamido)propyl, or 3-(tert-butoxycarbamido)propyl,which can be employed as N_(α)-protected amino acid derivatives inpeptide synthesis.

Another object of the present invention is to provide methods for thepreparation of said N_(α)-Nsc-amino acids. These and other objects ofthe present invention will be apparent from the following description

DETAILED DESCRIPTION OF THE INVENTION

N_(α)-Nsc-amino acids (I) of the present invention can be prepared bythe treatment of amino acids of the general formula II, wherein R₁ andR₂ represent the definitions meanings given for formula I, with2-(4-nitrophenylsulfonyl)ethylchloroformate III in mixed aqueous/organicsolvent in the presence of a base and at a temperature of from 0 to 40°C., preferably from 0 to 20° C. (Scheme 1).

Chloroformate III is introduced into the reaction in amounts of from 0.5to 1.5 molar equivalents, preferably from 0.7 to 0.9, as related to theamino acid. As an organic component of the solvent any aprotic organicsolvent may be used which is capable of dissolving the acylating reagentand is mixible with water, for example, acetonitrile, DMF,tetrahydrofuran or dioxane. A base may be an organic or inorganic base,for example, sodium or potassium carbonate, magnesium or calcium oxide,triethylamine, or N-methylmorpholine.

According to another method of the present invention, amino acids of thegeneral formula II are firstly converted into N,O-trimethylsilyldervatives using methods known in the art and then treated withequivalent amount of chloroformate III in anhydrous organic solvent, forexample, dichloromethane in the presence of base, for example, tertiaryamine. After aqueous hydrolysis of the intermediate acylatedtrimethylsilyl derivatives desirable N_(α)-Nsc-amino acids I areobtained in a free form.

Derivatives of the formula I, wherein R₁ is hydrogen, and R₂ representsN-(4,4′-dimethoxybenzhydryl) carboxamidomethyl or2-[N-(4,4′-dimethoxybenzhydryl)carboxamido]ethyl, may be prepared by thereaction of the known derivatives I, wherein R₁ is hydrogen, and R₂represents carboxamidomethyl or 2-(carboxamido)ethyl, with4,4′-dimethoxybenzhydrol in an organic solvent in the presence of astrong acid. As a solvent, acetic acid may be used, and as a strong acidsulfuric or methanesulfonic acid may be used.

A derivative of the formula I, wherein R₁ is hydrogen, and R₂ represents3-guanidinopropyl(N_(α)-Nsc-Arg-OH), may be prepared by the reaction ofarginine with 2-(4-nitrophenylthio) -ethylchloroformate in water in thepresence of sodium bicarbonate, resulting N_(α)-2-(4-nitrophenylthio)ethoxycarbonyl -arginine which is then oxidized intothe desired N_(α)-Nsc-arginine with hydrogen peroxide in an organicsolvent, for example, ethanol or acetic acid.

A derivative of the formula I, wherein R₁ is hydrogen, and R₂ representsimidazolyl-2-methyl(N_(α)-Nsc-His-OH), may be prepared fromN_(α)-Nsc-N_(im)-protected histidine derivative by selectivedeprotection of imidazole ring, for example, by selective acidicdetritylation of known N_(α)-Nsc-N_(im)-triphenylmethyl-histidine.

It is seen from the molecular formula that the compounds of formula Ihave an asymmetric α-carbon atom. Because the α-carbon atom does notparticipate in reactions employed for the preparation of compounds offormula I, the configuration of this chiral center existing in startingamino acids II is retained in the resulting N_(α)-Nsc-derivatives I.Therefore, it is clear that the methods of the present invention can beused for the preparation of N_(α)-Nsc-amino acids I in any chiral form(L or D), as well as racemic compounds, depending on the configurationof the compound II.

The meaning of R₁ and R₂ substituents in derivatives of the formula Iaccording to the present invention correspond to structures of sidechains of known α-amino acids containing or not containing protectivegroups known in the art (Table 1).

TABLE 1 Meanings of R₁ and R₂ substituents in compounds I No R1 R2 Aminoacid Abbreviation I-1 H Hydroxymethyl Serine Nsc-Ser-OH I-2 H1-Hydroxyethyl Threonine Nsc-Thr-OH I-3 H 4-Hydroxybenzyl TyrosineNsc-Tyr-OH I-4 H Imidazolyl-2-methyl Histidine Nsc-His-OH I-5 HBenzyloxymethyl O-Benzyl-serine Nsc-Ser(Bzl)-OH I-6 H 1-BenzyloxyethylO-Benzyl-threonine Nsc-Thr(Bzl)-OH I-7 H 4-BenzyloxybenzylO-Benzyl-tyrosine Nsc-Tyr(Bzl)-OH I-8 H Benzyloxy- Aspartic acid β-Nsc-Asp(OBzl)- carbonylmethyl benzyl-ester OH I-9 H 2-(Benzyloxy-Glutamic acid γ- Nsc-Glu(OBzl)- carbonyl)ethyl benzyl-ester OH I-10 HS-Benzylthiomethyl S-Benzyl-cysteine Nsc-Cys(Bzl)-OH I-11 H S-Diphenyl-S-Diphenylmethyl- Nsc-Cys(Dpm)- methyl)thiomethyl cysteine OH I-12 H4-Benzyloxy- Nε-Benzyloxy- Nsc-Lys(Z)-OH carbamido)-butylcarbonyl-lysine I-13 H 3-Guanidinopropyl Arginine Nsc-Arg-OH I-14 H3-(N^(G)-Toluene- N^(G)-Toluene- Nsc-Arg(Tos)-OH sulfonyl)guanidino-sulfonyl-arginine propyl I-15 H 3-(N^(G)- NO^(G)-Nitro-arginine Nsc-Nitro)guanidino- Arg(NO₂)—OH propyl I-16 H 3-[N^(G)-(4-Methoxy-N^(G)-(4-Methoxy- Nsc-Arg(Mtr)-OH 2,3,6-trimethyl- 2,3,6-trimethyl-benzenesulfonyl)] benzenesulfonyl)- guandinopropyl arginine I-17 HN-(4,4′-Dimethoxy- Nβ-(4,4′- Nsc-Asn(Mbh)- benzhydryl)car-Dimethoxybenzhy- OH boxamidomethyl dryl)-asparagine I-18 H 2-[n-(4,4′-Nγ-(4,4′- Nsc-Gln(Mbh)- Dimethoxy- Dimethoxybenzhy- OH benzhydryl)car-dryl)-glutamine boxamido]ethyl I-19 H S-tert-Butyl- S-tert-Butylthio-Nsc-Cys(StBu)- dithiomethyl cysteine OH I-20 H 4-[2-(4-Nitrophenyl-Nε-[2-(4-Nitro- Nsc-Lys(Nsc)-OH sulfonyl)ethoxy- phenylsulfonyl)-carbamido]butyl ethoxy carbonyl]- lysine I-21 H Propyl NorvalineNsc-Nva-OH I-22 H Butyl Norleucine Nsc-Nle-OH I-23 H Ethylα-Aminobutyric Nsc-Abu-OH acid I-24 H 3-(benzyloxy Nδ-Benzyloxy-Nsc-Om(Z)-OH carbamido)-propyl carbonyl-omitine I-25 H 3-(tert-Butoxy-Nδ-tert-Butoxy- Nsc-Om(Boc)- carbamido)-propyl carbonyl-omitine OH

Together with N_(α)-Nsc-amino acids disclosed in InternationalPublication No. WO96/25394, the compounds of the formula I shown inTable 1 may be used to implement a wide variety of tactical approachesto the peptide synthesis, including not only solid phase but alsosolution syntheses. Using N_(α)-Nsc-protection it is now possible todevelop synthetic schemes of peptide assembly in solution on the basisof tert-butyl type or benzyl type or a mixed type of side-chainprotection, and also to apply minimal side-chain protection tactics.

The present invention, therefore, is regarded as an addition to, and acontinuation of, aforementioned International Publication No.WO96/25394.

The invention will now be described by way of examples which areprovided as an illustration and are not intended as being limiting. Allof the amino acids in the following description have L-configurationunless otherwise indicated.

EXAMPLE 1 N_(α)-Nsc-O-Benzyl-Tyrosine (I-7)

8.15 g of O-benzyl-tyrosine and 7.7 g of potassium carbonate weredissolved in 150 ml of water-dioxane mixture (3:1, v/v) and cooled in anice bath, then a solution of 7.5 g of 2-(4-nitrophenylsulfonyl)ethylchloroformate III in 70 ml of dioxane was added dropwise within 15 minwith stirring. The cooling bath was removed and the mixture was stirredfor an additional 20 min, then evaporated to ca. 100 ml under reducedpressure and transferred into a separating funnel. 100 ml of water wasadded, and the resulting solution was extracted with 2×50 ml of ethylacetate. The aqueous layer was separated, acidified to pH 2 with 40%sulfuric acid with cooling in an ice bath, then extracted with 3×80 mlof ethyl acetate. Combined extracts were dried over anhydrous sodiumsulfate and evaporated at reduced pressure. Recrystallization of theresidue from ethyl acetate-hexane gave the desired product I-7 in a formof white crystalline powder (70%). For characterization see Table 2(Example 7).

EXAMPLE 2 N_(α)-Nsc-Threonine (I-2)

3.58 g of threonine and 100 ml anhydrous dichloromethane were placedinto 250 ml round-bottom flask equipped with a reflux condenser anddropping funnel. To the suspension, 9.7 ml of chlorotrimethylsilane wasadded with vigorous stirring, and the mixture was heated to boiling for1 hr. The resulting solution was cooled in an ice bath, then 9.0 ml oftriethylamine and 9.0 g of chloroformate III were added with stirring.The mixture was stirred for 20 min in an ice bath, then for anadditional 1.5 hr at room temperature. The solvent was evaporated atreduced pressure, and the residue was distributed between 200 ml ofethyl acetate and 250 ml of 2.5% aqueous sodium bicarbonate. The aqueouslayer was separated, washed with 50 ml of ether, acidified to pH 2 with1 N hydrochloric acid, then extracted with 3×70 ml of ethyl acetate. Thecombined extracts were dried with anhydrous sodium sulfate andevaporated at reduced pressure. Trituration of the residue withpetroleum ether gave the desired product I-2 in a form of white powder(75%). For characterization see Table 2 (Example 7).

EXAMPLE 3 N_(α)-Nsc-N_(δ)-Benzyloxycarbonyl-Ornitine (I-24)

8.0 g of N₆₇ -benzyloxycarbonyl-ornitine and 120 ml of anhydrousdichloromethane were placed into 250 ml round-bottom flask equipped witha reflux condenser and dropping funnel. To the mixture, 12.7 ml oftriethylamine and then 7 ml of chlorotrimethylsilane were added withvigorous stirring, and the mixture was heated to boiling for 1.5 hr. Thereaction mixture was then cooled in an ice bath, 9.0 g of chloroformateIII was added at once, and stirring was continued for 1.5 hr at roomtemperature. The solvent was evaporated at reduced pressure, and theresidue was distributed between 250 ml of ethyl acetate and 250 ml of2.5% aqueous sodium bicarbonate. The aqueous layer was separated, washedwith 50 ml of ether, acidified to pH 2 with 1 N hydrochloric acid, thenextracted with 2×100 ml of ethyl acetate. The combined extracts weredried with anhydrous sodium sulfate and evaporated at reduced pressure.Recrystallization of the residue from ethyl acetate-hexane gave thedesired product I-24 in a form of white powder (84%). Forcharacterization see Table 2 (Example 7).

EXAMPLE 4 N_(α)-Nsc-N(4,4′-Dimethoxybenzhydryl)-Asparagine (I-17)

3.89 g of N_(α)-Nsc-asparagine and 2.6 g of 4,4′-dimethoxybenzhydrolwere dissolved in 25 ml of glacial acetic acid. To the solution 0.4 mlof methanesulfonic acid was added, and the mixture was allowed to standovernight at room temperature. The resulting mixture was then pouredinto 100 ml of ice-cold water with mixing, the formed precipitate wasfiltered off, washed with water and then with ether. The crude productwas dissolved in 10 ml of warm DMF, filtered and reprecipitated withether. The precipitate was collected by filtration, washed with etherand dried in vacuo yielding the desired compound I-17 as a crystallinepower (80%). For characterization see Table 2 (Example 7).

EXAMPLE 5 N_(α)-Nsc-Arginine (I-1 3)

To the solution of 1.7 g of sodium bicarbonate and 4.22 g of argininehydrochloride in 25 ml of water stirred at room temperature 5.74 g of2-(4-nitrophenylthio)ethylchloro-formate was added in five equalportions during 30 min, then stirring was continued for additional 2hrs. The mixture was saponified to pH 8.5 with conc. ammonia solutionand left for 3 hrs at 4° C. The precipitate formed was collected, washedwith ice-cold water, dried on the filter with suction and dissolved in30 ml of acetic acid. To the solution, 4 ml of 34% hydrogen peroxide wasadded, and the mixture was left for 30 hrs at room temperature. Then themixture was evaporated to dryness, and the residue was triturated withethyl acetate, collected by filtration, washed with ether and dried onair to give the desired product I-13(88%). For characterization seeTable 2 (Example 7).

EXAMPLE 6 N_(α)-Nsc-Histidine (I-4)

6.6 g of N_(α)-Nsc-N_(im)-triphenylmethyl-histidine was dissolved in 30ml 90% aqueous acetic acid and boiled under reflux for 30 min. Aftercooling the precipitate was filtered off, and the filtrate wasevaporated. The residue was triturated with ether and collected byfiltration giving the desired compound I-4 in 95% yield. Forcharacterization see Table 2 (Example 7).

EXAMPLE 7 Properties of N_(α)-Nsc-amino acids I

Shown in Table 2 are the compounds of formula I which were preparedutilizing provided methods described in detail in examples 1-6. Figuresin the column “Method” correspond to numbers of examples whereparticular methods are described. Chromatographic mobility values R_(f)are shown for thin-layer chromatography sheets Alufolien Keselgel 60F₂₅₄ (Merck, Darmstadt, Germany); chloroform/methanol/acetic acid,95:5:3, (A) and benzene/acetone/acetic acid, 100:50:3, (B), were used asdeveloping solvents, spots were detected by UV-absorbance and/or byninhydrin reaction. Molecular ion masses (M+H)⁺ were measured usingMS-BC-1 time-of-flight mass spectrometer with Cf²⁵² radiation-promoteddesorption (Electrom SPA, Sumy, Ukraine).

TABLE 2 Properties of N_(α)-Nsc-amino acids I Molecular Molecular ion,(M + H)⁺ Entry Compound Method R_(f)(A) R_(f)(B) formula Calcd Found 1 23 4 5 6 7 8 I-1 Nsc-Ser-OH 2 0.40 0.30 C₁₂H₁₄N₂O₉S 363.32 363.6 I-2Nsc-Thr-OH 2 0.42 0.35 C₁₃H₁₆N₂O₉S 377.35 377.6 I-3 Nsc-Tyr-OH 2 0.400.37 C₁₈H₁₈N₂O₉S 439.44 439.9 I-4 Nsc-His-OH 6 0.15 0.10 C₁₅H₁₆N₄O₈S413.40 413.1 I-5 Nsc-Ser(Bzl)-OH 1,3 0.70 0.55 C₁₉H₂₀N₂O₉S 453.46 453.8I-6 Nsc-Thr(Bzl)-OH 3 0.70 0.55 C₂₀H₂₂N₂O₉S 467.49 467.9 I-7Nsc-Tyr(Bzl)-OH 1,3 0.75 0.55 C₂₅H₂₄N₂O₉S 529.57 530.1 I-8Nsc-Asp(OBzl)-OH 3 0.60 0.45 C₂₀H₂₀N₂O₁₀S 481.47 480.9 I-9Nsc-Asp(OBzl)-OH 3 0.62 0.43 C₂₁H₂₂N₂O₁₀S 459.50 495.1 I-10Nsc-Cys(Bzl)-OH 3 0.75 0.60 C₁₉H₂₀N₂O₈S₂ 469.53 469.3 I-11Nsc-Cys(Dpm)-OH 1,3 0.75 0.70 C₂₅H₂₄N₂O₈S₂ 545.63 545.2 I-12NsC-Lys(Z)-OH 3 0.65 0.50 C₂₃H₂₇N₃O₁₀S 538.58 538.3 I-13 Nsc-Arg-OH 50.10 0.05 C₁₅H₂₁N₅O₈S 432.45 432.8 I-14 Nsc-Arg(Tos)-OH 1,2 0.35 0.30C₂₂H₂₇N₅O₁₀S₂ 586.64 586.9 I-15 Nsc-Arg(NO₂—OH 2 0.25 0.15 C₁₅H₂₀N₆O₁₀S477.45 477.1 I-16 Nsc-Arg(Mtr)-OH 3 0.43 0.35 C₂₅H₃₃N₅O₁₀S₂ 628.73 629.4I-17 Nsc-Asn(Mbh)-OH 4 0.55 0.38 C₂₈H₂₉N₃O₁₁S 616.65 616.1 I-18Nsc-Gln(Mbh)-OH 4 0.58 0.33 C₂₉H₃₁N₃O₁₁S 630.68 630.2 I-19Nsc-Cys(StBu)-OH 3 0.60 0.45 C₁₆H₂₂N₂O₈S₃ 467.57 567.2 I-20Nsc-Lys(Nsc)-OH 3 0.55 0.40 C₂₄H₂₈N₄O₁₄S₂ 661.66 662.1 I-21 Nsc-Nva-OH 20.65 0.50 C₁₄H₁₈N₂O₈S 375.39 375.6 I-22 Nsc-Nle-OH 2 0.65 0.50C₁₅H₂₀N₂O₈S 389.42 389.6 I-23 Nsc-Abu-OH 2 0.60 0.47 C₁₃H₁₆N₂O₈S 375.40375.8 I-24 Nsc-Om(Z)-OH 3 0.62 0.47 C₂₂H₂₅N₃O₁₀S 533.55 534.0 I-25Nsc-Om(Boc)-OH 3 0.65 0.43 C₁₉H₂₇N₃O₁₀S 490.53 491.0

What is claimed is: 1.N_(α)-2-(4-nitrophenylsulfonyl)ethoxycarbonyl-amino acids having thegeneral formula:

wherein R₁ represents a hydrogen atom, and R₂ may representhydroxymethyl, 1-hydroxyethyl, 4-hydroxybenzyl, imidazolyl-2-methyl,benzyloxymethyl, 1-benzyloxyethyl, 4-benzyloxybenzyl,benzyloxycarbonylmethyl, 2-(benzyloxycarbonyl)ethyl, S-benzylthiomethyl,S-(diphenylmethyl)thiomethyl, 4-(benzyloxycarbamido)butyl,3-guanidinopropyl, 3-(N^(G)-toluenesulfonyl)guanidinopropyl,3-(N^(G)-nitro)guanidinopropyl,3{N^(G)(4-methoxy-2,3,6-trimethylbenzenesulfonyl)}-guanidinopropyl,N-(4,4′-dimethoxybenzhydryl)carboxamidomethyl,2-{N-(4,4′-dimethoxybenzhydryl)carboxamido}ethyl,S-tert-butyldithiomethyl,4-{2-(4-nitrophenylsulfonyl)ethoxycarbamido}-butyl, propyl, butyl, orethyl.
 2. A method for preparing N_(α)-2-(4-nitrophenylsulfonyl)ethoxycarbonyl amino acids according to claim 1 comprising reacting anamino acid of the general formula II HNR₁-CHR₂-COOH  II wherein R₁ andR₂ are the same as defined in claim 1, with 2-(4-nitrophenylsulfonyl)ethoxycarbonyl chloroformate in a mixed aqueous-organic solvent in thepresence of a base.
 3. A method for preparingN_(α)-2-(4-nitrophenylsulfonyl) ethoxycarbonyl amino acids according toclaim 1 comprising: a) converting an amino acid of the general formulaII HNR₁-CHR₂-COOH  II wherein R₁ and R₂ are the same as defined in claim1, into O,N-trimethylsilylated derivatives; b) reacting saidO,N-trimethylsilylated derivatives with 2-(4-nitrophenylsulfonyl)ethoxycarbonyl chloroformate in an aprotic solvent in the presence of abase, followed by hydrolysis.
 4. A method for preparingN_(α)-2-(4-nitrophenylsulfonyl) ethoxycarbonyl amino acids according toclaim 1 wherein R₁ represents a hydrogen atom, and R₂ representsN-(4,4′-dimethoxybenzhydryl)carboxamidomethyl or2-{N-(4,4′-dimethoxybenzhydryl)carboxamido}ethyl, comprising reactingN_(α)-2-(4-nitrophenylsulfonyl)-ethoxycarbonyl-asparagine orN_(α)-2-(4-nitrophenylsulfonyl)-ethoxycarbonyl-glutamine, with4-4′-dimethoxybenzhydrol in an organic solvent in the presence of anacid.
 5. A method for preparing N_(α)-2-(4-nitrophenylsulfonyl)ethoxycarbonyl amino acids according to claim 1, wherein R₁ represents ahydrogen atom, and R₂ represents 3-guanidinopropyl, comprising (a)reacting arginine with 2-(4-nitro-phenylthio) ethylchloroformate inwater in the presence of sodium bicarbonate; and (b) oxidizing the thusformed N_(α)-2-(4-nitrophenylthio) ethoxycarbonyl-arginine with hydrogenperoxide in an organic solvent.
 6. A method for preparingN_(α)-2-(4-nitrophenylsulfonyl) ethoxycarbonyl amino acids according toclaim 1 wherein R₁ represents a hydrogen atom, and R₂ representsimidazole-2-methyl, comprising reactingN_(α)-2-(4-nitrophenylsulfonyl)-ethoxycarbonyl-N_(im)-triphenylmethyl-histidinewith acid.